Bornheim L M, Grillo M P
Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California 94143-0450, USA.
Chem Res Toxicol. 1998 Oct;11(10):1209-16. doi: 10.1021/tx9800598.
Cannabidiol (CBD) is a major constituent of marijuana and a potent inhibitor of P450-mediated hepatic drug metabolism. Mouse P450 3A11 metabolism of [14C]CBD resulted in the formation of radiolabeled P450, which after digestion with lysyl endopeptidase C (Lys-C) and HPLC resolution of peptides, revealed one major broadly eluting peak of radioactivity. Electrophoresis/autoradiography of this peak identified several peptide bands, one of which was predominantly radiolabeled and had an apparent molecular mass of approximately 6 kDa. Amino-terminal sequence determination of this band revealed the presence of two peptides whose sequences identified them as Ala344-Lys379 and Gly426-Lys454. To characterize the reactive species that may be generated during P450 3A11-catalyzed CBD metabolism, reduced glutathione (GSH) was used as a trapping agent for possible electrophilic metabolites. Incubation of P450 3A11 in the presence of cofactors NADPH, CBD, and [3H]GSH resulted in the formation of a radiolabeled product which was absent in incubations lacking any of the cofactors. The UV absorption spectra of this compound indicated absorbances at approximately 220, 275, and 350 nm, and mass spectral analysis revealed prominent ions at m/z 634, 599, 505, 402, and 359, ions consistent with that of a GSH adduct of CBD-hydroxyquinone. A synthetic CBD-hydroxyquinone-GSH adduct was also prepared and had UV absorption and mass spectra nearly identical to that of the P450-mediated CBD-GSH adduct. CBD-hydroxyquinone formation may be the penultimate oxidative step involved in CBD-mediated modification and inactivation of P450 3A11.
大麻二酚（CBD）是大麻的主要成分，也是细胞色素P450介导的肝脏药物代谢的强效抑制剂。小鼠细胞色素P450 3A11对[14C]CBD的代谢导致放射性标记的细胞色素P450形成，在用赖氨酰内肽酶C（Lys-C）消化并通过高效液相色谱法分离肽段后，显示出一个主要的放射性宽泛洗脱峰。该峰的电泳/放射自显影片鉴定出几个肽条带，其中一个主要被放射性标记，表观分子量约为6 kDa。该条带的氨基末端序列测定显示存在两种肽，其序列将它们鉴定为Ala344-Lys379和Gly426-Lys454。为了表征在细胞色素P450 3A11催化的CBD代谢过程中可能产生的反应性物种，还原型谷胱甘肽（GSH）被用作可能的亲电代谢物的捕获剂。在辅因子NADPH、CBD和[3H]GSH存在下孵育细胞色素P450 3A11导致形成放射性标记产物，而在缺乏任何一种辅因子的孵育中则不存在该产物。该化合物的紫外吸收光谱表明在约220、275和350 nm处有吸收，质谱分析显示在m/z 634、599、505、402和359处有突出离子，这些离子与CBD-羟基醌的GSH加合物一致。还制备了一种合成的CBD-羟基醌-GSH加合物，其紫外吸收光谱和质谱与细胞色素P450介导的CBD-GSH加合物几乎相同。CBD-羟基醌的形成可能是CBD介导的细胞色素P450 3A11修饰和失活所涉及的倒数第二步氧化步骤。
